The alternative sigma factor c^ plays a central role in Salmonella pathogenesis. In the initial project period, we have found that Salmonella mutants lacking aE exhibit enhanced susceptibility to oxidative stress and antimicrobial peptides, reduced acid tolerance, deficient expression of virulence genes, impaired survival in phagocytic cells and markedly diminished virulence in mice. Furthermore we have identified novel OE- dependent loci, demonstrated that OE interacts with the other four alternative sigma factors in Salmonella, and discovered that aE complements the actions of the phage shock (psp) response in maintaining proton motive force. In this competing renewal application, our specific aims are to: [1] Comprehensively characterize the regulatory networks interconnecting the alternative sigma factors of Salmonella; [2] Analyze the mechanism of OE activation and its stimulation of Salmonella Pathogenicity Island 2 (SPI-2) gene expression in response to acid pH; [3] Examine the complementary actions of aE and PspA in promoting resistance to antimicrobial peptides and the maintenance of proton motive force; [4] Assess the roles of specific oE-dependent genes in resistance to acid pH, oxidative stress, antimicrobial peptides and virulence. Recent work in our laboratory and many others has revealed the enormous complexity and interconnectedness of bacterial regulatory networks. Through the proposed studies, we hope to provide a detailed mechanistic understanding of one of the most important regulatory pathways in Salmonella and to reveal novel insights into its physiologic functions.